Our research program focuses on the mechanisms of retroelement action. The most medically important class of these elements are the retroviruses, examples of these are responsible for AIDS as well as several types of tumor formation. Our approach to understanding the complex interactions between the element and its host is to study retrotransposons, a family of elements that are closely related to retroviruses. A significant advantage of studying retrotransposons is they exist in hosts such as yeast that can be studied using sophisticated molecular genetic techniques. The specific element we have chosen to study is the retrotransposon Tf1 of the fission yeast Schizosaccharomyces pombe. We have previously shown that Tf1 has several structural similarities to retroviruses including two long terminal repeats (LTRs) and coding sequences with significant homology to retroviral protease (PR), reverse transcriptase (RT), and integrase (IN). More importantly, we have developed an in vivo assay for transposition that we used to shown that Tf1 is functional and can transpose by integrating a new copy of itself into an unrelated section of the genome. Results from work done during our first nine months at NIH include significant improvements made to the in vivo transposition assay. The assay depends on the overexpression of Tf1 transcripts containing a selectable marker that is reverse transcribed during transposition and inserted into the host genome. By modifying conditions we increased our previously reported transposition frequency of 0.5%/(11 cell generations) 40-fold to 18%. This improvement allowed us to develop a new transposition assay that is now being used to screen large numbers of cells for mutations that alter the transposition frequency. Our biochemical analysis of the transposon proteins has included the use of immunoprecipitation and in vivo protein labeling to determine the order and identity of the protease cleavage events of the 150 kDa precursor protein that results in the production of the mature Tf1 proteins.